A feature of certain acute lymphoblastic leukemis (ALLs) is the sequence “evolution” of clonotypes associated with the disease. Although treatment outcomes in childhood acute lymphoblastic leukemias (ALL) have improved dramatically over the past fifty years, 10 to 15% of patients will ultimately relapse, usually with disease that is highly refractory to additional therapy, e.g. Pui et al, Lancet Oncol., 2:597-607 (2001); Pui et al, New Engl. J. Med., 350: 1535-1548 (2004); Tallen et al, J. Clin. Oncol., 28; 2339-2347 (2010). It is generally thought that these relapses are due to residual leukemic cells that are resistant to therapy and remain undetected during clinical remission. Several potential biological explanations tor the persistence of leukemic populations have been proposed, including the presence of multiple clones at diagnosis with varying susceptibilities to cytotoxic drugs or the outgrowth of resistant clones during treatment, e.g. Rosenquist et al, Eur. J. Haematol., 63: 171-179(1999).
Clonality of B-cell populations can be assessed by analysis of gene rearrangements that occur at the immunoglobulin heavy chain (IgH) gene locus. Early in B-cell development, somatic recombination at the IgH gene locus gives rise to unique rearrangements of the variable (VH), diversity (D), and joining (JH) gene segments, e.g. Tonegawa, Nature, 302: 575-581 (1983); Alt et al, Immunol. Today, 13: 306-314 (1992). In this two-step process, recombination signal sequences mediate D to JH joining, which is followed by VH to D-JH joining, e.g. Alt et al, EMBO J., 3: 1209-1219 (1984); Hiom et al, Cell, 88: 65-72 (1997); Hess et al, Genes Dev., 3: 1053-1061 (1989). During this recombination, non-templated nucleotides (N-bases) may be added at the junctions between gene segments, and other nucleotides may be deleted from the VH, D, and JH germline sequences, Alt et al, Proc. Natl. Acad. Sci., 79: 4118-4122 (1982). The resulting unique VHDJH rearrangements are used as clonotypic markers in precursor-B-cell ALL.
Precursor-B-cell ALL is generally thought to be a clonal disease resulting from malignant transformation and expansion of a single B-cell, e.g. Steenbergen et al, Leukemia, 1 I: 1258-1265 (1997). PCR-based methods have, however, shown changes in clonal IgH rearrangements between initial diagnosis and relapse in a significant proportion of pre-B ALL cases, Beishuizen et al Blood, 83: 2238-2247 (1994); Li et al, Leukemia Research, 25: 1033-1045 (2001); Szczepanski et al, Blood, 99: 2315-2323 (2002). These changes at the IgH locus could represent the persistence of ancestral clones that later expand, or continued evolution of a. primary ancestral clone its the setting of genotoxic anti-neoplastic therapy.
Current methods of monitoring leukemia clonotypes are not well suited for detecting changes or evolution of the sequences of leukemic clonotypes. It would be highly advantageous for patterns suffering from such diseases if there were available methods for detecting and monitoring sequence changes or evolution of leukemic clonotypes.